The present invention relates to an apparatus for electrically contacting biological cells suspended in a liquid, and more particularly to an apparatus comprising a substrate having at least one opening, a means for immobilizing a cell on the opening, and at least one electrode for electrically contacting the immobilized cell.
The invention further relates to a method of electrically contacting biological cells suspended in a liquid, comprising the steps of immobilizing a cell above an opening provided in a substrate, and contacting the immobilized cell via at least one electrode.
An apparatus known from DE 197 12 309 A1 (which is equivalent to U.S. Pat. No. 6,315,940 assigned to the present assignee and which is incorporated here by reference) comprises a microelectrode arrangement in which the cells are captured in microcuvettes at the bottom of which an electrode is located. The electrode is provided with a central suction channel in which a low pressure can be generated by connecting channels which run underneath the electrodes. Thus, it is possible to attract individual cells selectively to the electrodes and immobilize them on the electrodes with a certain contact pressure. It is feasible to use the known apparatus for measurements on cells, but only from the outside of their cell membranes.
Microelectrode arrangements of this kind are generally used for studying biological cells. The microelectrode arrangements serve, for example, to stimulate the cells or to pick up cell potentials. The studies may be carried out in a biological environment or in an artificial environment. In conventional microelectrode arrangements, however, it depends more or less on chance whether or not one or another cell settles on a particular electrode. Furthermore, the cells generally settle on an electrode only with partial coverage, so that stimulating the cell or picking up a cell potential is limited to this partial area.
Although the apparatus known from DE 197 12 309 A1 already mentioned above removes most of these disadvantages, in particular by ensuring that only one cell settles on any micro-electrode, it is nevertheless still a matter of chance as to whether a cell and which cell is captured in the microcuvette. Furthermore, it cannot be ruled out that particles present in the liquid are sucked into the microcuvette, which do allow a measurement to be carried out, although its results are unusable.
Another characteristic of the known apparatus and the known method is the fact that measurements can be carried out only on the outside of the cell membranes. Intracellular measurements are not possible.
Intracellular measurements, however, can be carried out using the well known “patch clamp” technique in which a “membrane patch clamp” is used; see, for example, Neher and Sakmann: “Die Erforschung von Zellsignalen mit der Patch-Clamp-Technik”, Spektrum der Wissenschaften, May 1992, p. 48.
In this technique, a micropipette filled with electrically conductive liquid is selectively brought close to an adherent cell and is lowered down onto the membrane thereof which, as a result, bulges slightly inward. The membrane patch enclosed by the pipette tip is then sucked in slightly by means of low pressure, thereby sealing off and electrically isolating the membrane patch from the surrounding liquid. This isolation is often referred to as gigaseal.
In this way it is possible to carry out, for example, measurements of ion channels on the outside of a membrane.
However, if the sealed-off membrane is sucked in further, the membrane patch is perforated so that there is access through the pipette orifice to the cell interior, which is sealed off hydrodynamically and electrically from the surrounding liquid, making intracellular measurements and stimulations possible (“whole cell patch”).
However, the patch clamp technique can only be carried out on adherent or otherwise immobilized cells, which the fragile glass pipette has to be brought close to by means of a micro-manipulator in each case. For this reason, the number of simultaneously contactable cells is extremely limited so that a plurality of cells can be processed only sequentially.
Therefore, this method is unsuitable for mass studies, as would be required, for example, in the area of drug screening, substance screening, etc.
Another disadvantage of the conventional patch clamp technique is the fact that it cannot be automated but is carried out manually by workers who need to have a lot of experience and manual skills.
From DE 198 27 957 A1, it is know to immobilize a cell on a glass support whose surface has a ring structure-like profile in order to improve cell adhesion. A conically projecting electrode whose sharp, ring-shaped tip cuts into the cell membrane is arranged on the glass support in the cell-supporting area. The known apparatus, however, has the disadvantage that the electrical sealing on the ring-shaped electrode tip is determined only by the “adhesive force” of the cell on the profiled surface.